This invention relates to a process for producing a maltooligosaccharide derivative very useful, for example, as a precursor of a maltooligosaccharide derivative having modified groups at the 6-position or the 4- and 6-positions of non-reducing end glucose unit which can be effectively used as a substrate for measuring .alpha.-amylase activity.
Measurement of .alpha.-amylase activity in samples, in particular, saliva, pancreatic juice, blood and urine in a human body gives an important indication for medical diagnoses. For example, in the case of pancreatitis, pancreas cancer and parotiditis, the level of .alpha.-amylase activity in blood and urine is much higher than its usual level. For the measurement, there has come to be generally employed in recent years a coupling enzyme method using as substrate a maltooligosaccharide derivative having a color-producing group at the reducing end and a non-reducing end glucose unit modified at the 6-position or the 4- and 6-positions.
In the synthesis of such a substrate having a non-reducing end glucose unit modified only at the 6-position or the 4- and 6-positions, if there is a common intermediate which permits free and easy introduction of various modifying groups only into the 6-position or the 4- and 6-positions of the non-reducing end glucose unit, it becomes possible to synthesize rapidly and efficiently a substrate for measuring .alpha.-amylase activity or an intermediate thereof, which has higher performance characteristics and a newer function. Therefore, the production and development of a substrate for measuring .alpha.-amylase activity proceed rapidly, and hence the common intermediate is of high utility value.
An example of the common intermediate is a polyacyl maltooligosaccharide derivative retaining the hydroxyl groups at the 4- and 6-positions of non-reducing end glucose unit. Employment of this intermediate permits selective and easy synthesis of a substrate having a non-reducing end glucose unit modified only in the hydroxyl group at the 6-position and a substrate having a non-reducing end glucose unit modified at the 4- and 6-positions with bridge type blocking groups. Moreover, since this intermediate is formed by acylation in all hydroxyl groups other than the hydroxyl groups at the 4- and 6-positions, it is highly lipophilic and is soluble in various organic solvents. Therefore, it has many.advantages, for example, in that the determination of reaction reagents, reaction conditions and the like is easy.
As heretofore known processes for producing a polyacyl maltooligosaccharide derivative retaining the hydroxyl groups at the 4- and 6-positions, there are processes disclosed in Japanese Patent Unexamined Publication Nos. 60-54395 (U.S. Pat. Nos. 4,709,020, 4,818,692) and 2-49796 (U.S. Pat. No. 5,011,923), etc., namely, a process which uses as starting material a commercially available oligosaccharide derivative having an optically measurable group at the reducing end and comprises blocking the 4- and 6-positions of non-reducing end glucose unit with a cyclic ketal (or acetal) type blocking group such as ethylidene group, isopropylidene group, benzylidene group or the like, acylating the thus treated derivative, and then deblocking the 4- and 6-positions. This method, however, is disadvantageous in that both the blocking step and the deblocking step require severe conditions and a troublesome procedure and that the yield is low.
The reasons for the low yield in the case of the blocking with the aforesaid cyclic ketal (or acetal) type blocking groups are, for example, as follows. The introduction of the blocking groups is naturally not easy. Because of the severe conditions in both the blocking step and the deblocking step (particularly in the deblocking step), some of modifying groups at the non-reducing end are removed. Also in positions other than the 4- and 6-positions of non-reducing end glucose unit, blocking by crosslinking takes place in the same glucose unit or between two glucose units (Carbohyd. Res., 185, 91-104 (1989)).
In addition to the above process, there is the process disclosed in the specification of EP 0512808 in which the present inventors have applied for a patent. This process also cannot give a sufficient yield because a considerable amount of polyacyl maltooligosaccharide derivatives retaining the hydroxyl groups at positions other than the 4- and 6-positions are produced as by-products together with a polyacyl maltooligosaccharide derivative retaining the hydroxyl groups only at the 4- and 6-positions.
Accordingly, there is an eager desire for the advent of a process for producing a polyacyl maltooligosaccharide derivative retaining the hydroxyl groups only at the 4- and 6-positions of non-reducing end glucose unit, easily in high yield under mild conditions by using as starting material a commercially available oligosaccharide derivative having an optically measurable group at the reducing end.